We are studying the secretin-VIP family of G-protein coupled receptors. This recently identified group of receptors has virtually no amino acid sequence conservation with the remaining majority of G-protein coupled receptors (the rhodopsin family). Much less is known about structure-function relationships and effector system coupling of the secretin-VIP family receptors. Their cloning has also raised new questions about their physiological roles and created tools for study of their distribution, regulation and pharmacology. We are currently pursuing the physiological role of the PTH2 receptor which we discovered 2 years ago. The receptor is most abundant in the brain but we detect no PTH, its only known ligand, in the CNS. We have now demonstrated the existence of a new neuropeptide, selective for the PTH2 receptor in hypothalamic extracts. We continue to work on the purification of that peptide and have developed a number of effective chromatography procedures. We now plan a large scale up. The PTH2 and PTH/PTHrP receptors are both activated by PTH but only the PTH/PTHrP receptor recognizes PTHrP. Since the receptors are quite similar in sequence we have been able to make functional chimeras between the two receptors and begin identifying the domains responsible for ligand selectivity. We have shown that at least two distinct domains of the PTH2 receptor differentiate between PTH and PTHrP and that a third, as yet unidentified region of the PTH/PTHrP receptor must participate in high affinity PTHrP binding. These data suggest that endogenous peptide ligands for these receptors bind to a large distributed surface of the receptor's extracellular domains. Rigorous pharmacological studies of peptide receptors have generally been limited by the lack of antagonists. Studies of PTH recognizing receptors suffer from relatively high background when membrane binding assays are performed. We have developed new assay conditions for ligand binding to PTH receptors addressing both of these issues, which allow quantitative evaluation of binding models. We have developed an antibody which selectively recognizes the PTH2 receptor and have begun using it to study the distribution and regulation of PTH2 receptor protein.